Transgenic technology is widely used in biotechnology. In eukaryotes, the temporal and spatial expression of transgenes is regulated by transcription factors through their interaction with enhancer elements. However, eukaryotic enhancer-promoter interactions lack the specificity for precise temporal and spatial patterns of transgenic expression. The expression of a transgene can be affected by (a) the promoter of a selectable marker gene that is closely linked with the transgene, and/or (b) by chromosomal genes that flank the transgene, which is often referred to as the “position effect”. These effects are often not desirable, especially when tissue-specific, precisely controlled, or optimal transgene expression is wanted.
It is known that a eukaryotic genome has organizational properties that rely on the ability of the chromosome to establish autonomous functional units. The polynucleotide sequences that separate these domains are called genetic insulator elements. These genetic insulator elements can buffer a transgene from position effects, so that an introduced transgene can be expressed independent of its location in the chromosome. In addition, a genetic insulator may repress nonspecific interactions between enhancers and promoters. Thus, it could be possible to obtain precise gene expression by using an appropriate genetic insulator to shield the effects of neighboring gene promoters.
Genetic insulators in fruit fly (Drosophila) include specialized chromatin structures (scs and scs′), which consist of 350 base pairs (bp) and 200 bp, respectively. These sequences are associated with chromatin structures and serve as boundaries that can prevent activation by enhancer elements. Similarly, genetic insulators are known in the chicken in the form of lysozyme “A” element and the β-globin LCR (HS4), which contain 242 bp. These insulators generally comprise 200-250 bp and function directionally. Additionally, the gypsy chromatin insulator of Drosophila (originally isolated from the gypsy retrotransposon) protects a gene and its regulatory elements from both positive and negative position effects (see, U.S. Pat. No. 6,229,070, incorporated by reference).
By contrast, there previously have been no genetic insulators isolated from plants. There is a need in the art to develop a plant genetic insulator to control transgene expression in plants.